Background: Azadirachta indica (Neem) contains bioactive triterpenoids such as nimbolide with anticancer potential, but poor solubility and bioavailability limit its use. We developed PLGA-based Neem extract nanoparticles (NE-PLGA NPs) to improve delivery, cytotoxicity, and in vivo efficacy against breast cancer.

Methods: Neem leaves were extracted with 80% ethanol, and NE-PLGA NPs were prepared by nanoprecipitation (extract:polymer 1:5 w/w). Nanoparticles averaged 128 ± 9 nm with 78% encapsulation efficiency and sustained release over 72 h. Cytotoxicity was assessed in MCF-7, 4T1, and A549 cells versus human dermal fibroblasts. In vivo efficacy was tested in BALB/c mice bearing orthotopic 4T1 tumors (n = 10/group), comparing vehicle, crude extract, NE-PLGA NPs, doxorubicin, and combination therapy. Tumor growth, metastasis, survival, and histological markers of proliferation, apoptosis, and angiogenesis were evaluated.

Results: NE-PLGA NPs significantly improved in vitro potency with IC50 values of 4.8–8.5 µg/mL compared to 22–34 µg/mL for crude extract, while maintaining >85% fibroblast viability at 50 µg/mL. Synergy with doxorubicin (CI 0.44–0.67) enhanced cytotoxicity. In vivo, NE-PLGA NPs reduced tumor volume by 68% and prolonged median survival from 28 to 46 days (p < 0.01). Combination therapy achieved 89% tumor inhibition, suppressed lung metastasis by 94%, and yielded 80% long-term survival beyond 60 days. Immunohistochemistry confirmed marked apoptosis (↑caspase-3, ↑TUNEL), reduced proliferation (↓Ki-67), and suppressed angiogenesis (↓VEGF). No hepatotoxicity, nephrotoxicity, or body weight loss was observed.

Conclusion: Neem-derived PLGA nanoparticles significantly potentiate antitumor activity, overcome resistance, and synergize with doxorubicin while remaining safe in vivo. NE-PLGA NPs represent a promising nanomedicine platform for future preclinical and translational anticancer development.